Abstract As genomic databases expand and artificial intelligence tools advance, there is a growing demand for efficient characterization of large numbers of proteins. To this end, here we describe a ...generalizable pipeline for high-throughput protein purification using small-scale expression in E. coli and an affordable liquid-handling robot. This low-cost platform enables the purification of 96 proteins in parallel with minimal waste and is scalable for processing hundreds of proteins weekly per user. We demonstrate the performance of this method with the expression and purification of the leading poly(ethylene terephthalate) hydrolases reported in the literature. Replicate experiments demonstrated reproducibility and enzyme purity and yields (up to 400 µg) sufficient for comprehensive analyses of both thermostability and activity, generating a standardized benchmark dataset for comparing these plastic-degrading enzymes. The cost-effectiveness and ease of implementation of this platform render it broadly applicable to diverse protein characterization challenges in the biological sciences.
To determine whether TMEM106B single nucleotide polymorphisms (SNPs) are associated with frontotemporal lobar degeneration (FTLD) in patients with and without mutations in progranulin (GRN) and to ...determine whether TMEM106B modulates GRN expression.
We performed a case-control study of 3 SNPs in TMEM106B in 482 patients with clinical and 80 patients with pathologic FTLD-TAR DNA-binding protein 43 without GRN mutations, 78 patients with FTLD with GRN mutations, and 822 controls. Association analysis of TMEM106B with GRN plasma levels was performed in 1,013 controls and TMEM106B and GRN mRNA expression levels were correlated in peripheral blood samples from 33 patients with FTLD and 150 controls.
In our complete FTLD patient cohort, nominal significance was identified for 2 TMEM106B SNPs (top SNP rs1990622, p(allelic) = 0.036). However, the most significant association with risk of FTLD was observed in the subgroup of GRN mutation carriers compared to controls (corrected p(allelic) = 0.0009), where there was a highly significant decrease in the frequency of homozygote carriers of the minor alleles of all TMEM106B SNPs (top SNP rs1990622, CC genotype frequency 2.6% vs 19.1%, corrected p(recessive) = 0.009). We further identified a significant association of TMEM106B SNPs with plasma GRN levels in controls (top SNP rs1990622, corrected p = 0.002) and in peripheral blood samples a highly significant correlation was observed between TMEM106B and GRN mRNA expression in patients with FTLD (r = -0.63, p = 7.7 × 10(-5)) and controls (r = -0.49, p = 2.2 × 10(-10)).
In our study, TMEM106B SNPs significantly reduced the disease penetrance in patients with GRN mutations, potentially by modulating GRN levels. These findings hold promise for the development of future protective therapies for FTLD.
Performance losses and aging mechanisms are investigated in state‐of‐the‐art PTB7:PC70BM solar cells. Inverted devices incorporating a vanadium pentoxide (V2O5) top contact have efficiencies of 8%. ...After aging the unencapsulated devices, no changes are observed in the open circuit voltage (Voc) or short circuit current (Jsc); however, the fill factor (FF) drops from 0.7 to 0.61. An s‐shape initially appears in the J–V curve after aging, which can be reduced by cycling through the J–V curve under illumination. This is discussed in context of the redox properties of V2O5. With impedance spectroscopy, it is demonstrated that changes to the contact interfaces are completely reversible and not responsible for the performance loss. Intensity modulated photocurrent spectroscopy combined with device modeling reveals that the loss in FF is due to trap formation in the active layer. Additionally it is observed that the performance of pristine devices is limited by optical absorption in the thin active layer and the build‐up of space charge which hinders carrier extraction.
Air stable PTB7:PC70BM solar cells with efficiencies of 8% are demonstrated. The device employs a V2O5 anodic transport layer. Impedance spectroscopy is applied to confirm that the device contact interfaces in this architecture are stable under ambient conditions. Intensity modulated photocurrent spectroscopy is used to model the optoelectronic response of solar cell active layer as a function of aging.
Solar cells based on conjugated polymer and fullerene blends have been developed as a low-cost alternative to silicon. For efficient solar cells, electron-hole pairs must separate into free mobile ...charges that can be extracted in high yield. We still lack good understanding of how, why and when carriers separate against the Coulomb attraction. Here we visualize the charge separation process in bulk heterojunction solar cells by directly measuring charge carrier drift in a polymer:fullerene blend with ultrafast time resolution. We show that initially only closely separated (<1 nm) charge pairs are created and they separate by several nanometres during the first several picoseconds. Charge pairs overcome Coulomb attraction and form free carriers on a subnanosecond time scale. Numerical simulations complementing the experimental data show that fast three-dimensional charge diffusion within an energetically disordered medium, increasing the entropy of the system, is sufficient to drive the charge separation process.
A period of elevated surface concentrations of airborne particulate matter (PM) in the UK in spring 2014 was widely associated in the UK media with a Saharan dust plume. This might have led to ...over-emphasis on a natural phenomenon and consequently to a missed opportunity to inform the public and provide robust evidence for policy-makers about the observed characteristics and causes of this pollution event. In this work, the EMEP4UK regional atmospheric chemistry transport model (ACTM) was used in conjunction with speciated PM measurements to investigate the sources and long-range transport (including vertical) processes contributing to the chemical components of the elevated surface PM. It is shown that the elevated PM during this period was mainly driven by ammonium nitrate, much of which was derived from emissions outside the UK. In the early part of the episode, Saharan dust remained aloft above the UK; we show that a significant contribution of Saharan dust at surface level was restricted only to the latter part of the elevated PM period and to a relatively small geographic area in the southern part of the UK. The analyses presented in this paper illustrate the capability of advanced ACTMs, corroborated with chemically-speciated measurements, to identify the underlying causes of complex PM air pollution episodes. Specifically, the analyses highlight the substantial contribution of secondary inorganic ammonium nitrate PM, with agricultural ammonia emissions in continental Europe presenting a major driver. The findings suggest that more emphasis on reducing emissions in Europe would have marked benefits in reducing episodic PM2.5 concentrations in the UK.
Small-scale bioreactors that are affordable and accessible would be of major benefit to the research community. In previous work, an open-source, automated bioreactor system was designed to operate ...up to the 30 mL scale with online optical monitoring, stirring, and temperature control, and this system, dubbed Chi.Bio, is now commercially available at a cost that is typically 1–2 orders of magnitude less than commercial bioreactors. In this work, we further expand the capabilities of the Chi.Bio system by enabling continuous pH monitoring and control through hardware and software modifications. For hardware modifications, we sourced low-cost, commercial pH circuits and made straightforward modifications to the Chi.Bio head plate to enable continuous pH monitoring. For software integration, we introduced closed-loop feedback control of the pH measured inside the Chi.Bio reactors and integrated a pH-control module into the existing Chi.Bio user interface. We demonstrated the utility of pH control through the small-scale depolymerization of the synthetic polyester, poly(ethylene terephthalate) (PET), using a benchmark cutinase enzyme, and compared this to 250 mL bioreactor hydrolysis reactions. The results in terms of PET conversion and rate, measured both by base addition and product release profiles, are statistically equivalent, with the Chi.Bio system allowing for a 20-fold reduction of purified enzyme required relative to the 250 mL bioreactor setup. Through inexpensive modifications, the ability to conduct pH control in Chi.Bio reactors widens the potential slate of biochemical reactions and biological cultivations for study in this system, and may also be adapted for use in other bioreactor platforms.
Aim
To conduct a retrospective population‐based study to examine the risk of developing diabetes after delivery in First Nations and non‐First Nations women in Alberta.
Methods
Delivery records ...(1999–2014) were linked to provincial administrative data, which allowed for a maximum follow‐up of 16 years after delivery. Prevalence of pregnancy risk factors were compared by First Nations status. Hazard ratios for diabetes after delivery by First Nations status, high pre‐pregnancy body weight (≥91 kg) and gestational diabetes status were estimated using the Cox proportional hazards model.
Results
Age‐adjusted prevalence of gestational diabetes (7.9% vs 4.6%; P<0.0001), high pre‐pregnancy body weight (18.8% vs 10.2%; P<0.0001) and diabetes after delivery (3.9% vs 1.1%; P<0.0001) were higher in First Nations women than in non‐First Nations women. Development of diabetes after delivery was higher with First Nations status (hazard ratio 3.0, 95% CI 2.6–3.4), high pre‐pregnancy body weight (hazard ratio 3.6, 95% CI 3.3–4.0) and gestational diabetes status (hazard ratio 19.2, 95% CI 17.9–20.6). The highest risk was within First Nations women with high pre‐pregnancy body weight and gestational diabetes (hazard ratio 54.8, 95% CI 45.2–66.5) compared to women without these three risk factors. Reduced prenatal visits per pregnancy (8.4 vs 10.7; P<0.0001) and delayed first prenatal visit (time to delivery 23.7 vs 26.7 weeks; P<0.0001) were observed in First Nations women compared to non‐First Nations women.
Conclusion
First Nations women are at greater risk of developing diabetes after pregnancy, with gestational diabetes being the strongest predictor. Strategies that target the specific needs of First Nations women before, during and after pregnancy are required.
Rapid diagnosis and appropriate antimicrobial therapy are of major importance to decrease morbidity and mortality in patients with blood stream infections (BSI). Blood culture, the current gold ...standard for detecting bacteria in blood, requires at least 24-48 hours and has limited sensitivity if obtained during antibiotic treatment of the patient. The aim of this prospective multicenter study was to clinically evaluate the application of a commercial universal 16S/18S rDNA PCR, SepsiTest™ (PCR-ST), directly on whole blood.
In total 236 samples from 166 patients with suspected sepsis were included in the study. PCR-ST results were compared to blood culture, the current gold standard for detecting BSI. Because blood cultures can give false-negative results, we performed an additional analysis to interpret the likelihood of bloodstream infection by using an evaluation based on clinical diagnosis, other diagnostic tests and laboratory parameters.
Clinical interpretation of results defined the detected organism to be contaminants in 22 of 43 positive blood cultures (51.2 %) and 21 of 47 positive PCR-ST results (44.7 %). Excluding these contaminants resulted in an overall sensitivity and specificity of the PCR-ST of 66.7 and 94.4 % respectively. Of the 36 clinically relevant samples, 11 BSI were detected with both techniques, 15 BSI were detected with PCR-ST only and 10 with blood culture only. Therefore, in this study, SepsiTest™ detected an additional 71 % BSI compared to blood culture alone.
More clinically relevant BSI were diagnosed by molecular detection, which might influence patient treatment. An improved SepsiTest™ assay suited for routine use can have additional value to blood culture in diagnosing bacteremia in septic patients.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
In accordance with worldwide data, the Robert Koch Institute (RKI) has reported a constant increase of syphilis cases in Germany over the past decade.
We analysed the data of all patients, referred ...to a Department of Ophthalmology in a tertiary referral centre in Düsseldorf, Germany between 2008 and 2019, who were tested for syphilis. The epidemiologic, demographic, clinical, diagnostic and therapeutic data were retrieved from the records and evaluated in a retrospective, descriptive, non-comparative study.
Syphilis serology was positive in 32/1840 (1.7%) patients, and was evenly distributed over this period. 26 (81.3%) were male, 19 (59.4%) belonged to a risk group. Ocular syphilis was the primary diagnosis for 29 patients (90.6%). The most frequent manifestation was uveitis (n = 20, 62.5%). By the end of therapy, 19 patients (59.4%) had an improved visual acuity.
The incidence of ocular syphilis cases has remained stable over the last decade.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Background. Pneumonia is the leading cause of death in children globally. Clinical algorithms remain suboptimal for distinguishing severe pneumonia from other causes of respiratory distress such as ...malaria or distinguishing bacterial pneumonia and pneumonia from others causes, such as viruses. Molecular tools could improve diagnosis and management. Methods. We conducted a mass spectrometry–based proteomic study to identify and validate markers of severity in 390 Gambian children with pneumonia (n = 204) and age-, sex-, and neighborhood-matched controls (n = 186). Independent validation was conducted in 293 Kenyan children with respiratory distress (238 with pneumonia, 41 with Plasmodium falciparum malaria, and 14 with both). Predictive value was estimated by the area under the receiver operating characteristic curve (AUC). Results. Lipocalin 2 (Lpc-2) was the best protein biomarker of severe pneumonia (AUC, 0.71 95% confidence interval, .64–.79) and highly predictive of bacteremia (78% 64%–92%), pneumococcal bacteremia (84% 71%–98%), and "probable bacterial etiology" (91% 84%—98%). These results were validated in Kenyan children with severe malaria and respiratory distress who also met the World Health Organization definition of pneumonia. The combination of Lpc-2 and haptoglobin distinguished bacterial versus malaria origin of respiratory distress with high sensitivity and specificity in Gambian children (AUC, 99% 95% confidence interval, 99%–100%) and Kenyan children (82% 74%–91%). Conclusions. Lpc-2 and haptoglobin can help discriminate the etiology of clinically defined pneumonia and could be used to improve clinical management. These biomarkers should be further evaluated in prospective clinical studies.
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